A Light Aggregometry Based Method To Measure The Percentage Of An Individual’s Platelets That Are Inhibited By Aspirin

Ex-vivo analysis of platelet inhibition by aspirin (ASA) suggests that an optimal decrease in platelet function occurs when 90-95% of platelets are inhibited with ASA. In patients with known vascular disease a daily aspirin decreases the probability of a future vascular event by about 25%. However, platelets from patients with diabetes mellitus (DM) are highly reactive with an increased turnover rate, and prophylaxis with a daily aspirin is not nearly as effective. Because platelet response among individuals is highly variable, we developed a light aggregation based assay that measures platelet function with 100% blockade of an individual’s platelet thromboxane A₂ receptor (100% “aspirinated” platelets). This assay is utilized to determine the percentage of platelets inhibited by aspirin, “aspirinated” platelets (ASA-PLTS).

Platelet prostaglandin agonist (PPA), (Analytical Control Systems, Fishers IN) an agonist known to be sensitive to the amount of platelet inhibition with aspirin, was used to develop a light aggregometry based technique that measures ASA-PLTS. A specific inhibitor of the thromboxane A₂ receptor SQ 29,548(S.Q.)(Cayman Chemicals, Ann Arbor, Mi.) was used to verify that maximal (100%) inhibition of platelets by aspirin was present 2 hrs after ingestion of 325mg ASA. In separate experiments platelets obtained 2 hrs after 325mg ASA (100% ASA-PLTS) were mixed with platelets from the same subject obtained just before aspirin ingestion (0% ASA-PLTS) so that the PPA light aggregation slopes could be measured with 95%, 90%, 85% and 80% ASA-PLTS. The slope of the PPA-AGG curve measured 24 hours after 325mg ASA was compared to the slopes obtained with known concentrations of ASA-PLTS using the formula T24-T2/ T0-T2. (T24= slope 24 hrs post ASA; T2=slope 2 hrs post ASA; T0=slope off ASA).

Two hours after ingestion of 325mg ASA the slopes of the PPA-AGG curves with and without S.Q. were identical in 7 normal subjects reflecting the aspirin response with 100% ASA-PLTS.

Because of the large range of response among individuals, eac subject served as their own control to develop an aspirin response curve for the different percentages of ASA-PLTS. 24 hours after 325mg ASA the percentage of ASA-PLTS in 6 of 7 normal subjects was greater than 90%.

1. The slope of the PPA stimulated light aggregation curve with and without S.Q. obtained 2 hrs after 325mg of aspirin in normal subjects can be used as a measure of 100% ASA-PLTS response. 2. Platelet response in most normal subjects 24 hours after aspirin shows that at least 90% of their platelets are “aspirinated”.

Comment

This technique can identify patients, like those with DM whose percentage of ASA-PLTS may fall below 90% at the end of the dosing interval. A shorter aspirin dosing interval in these patients may result in a higher percentage of ASA-PLTS at the end of the dosing period and possibly decrease the patient’s probability of a subsequent vascular event, but these questions can only be answered with clinical studies.

No relevant conflicts of interest to declare.